In general, organ function is dependent on an adequate supply of energy which is generated within the cell and can be converted into cellular work. Cellular metabolism generates adenosine triphosphate (ATP), a molecule which contains high energy bonds. It is the energy contained in the ATP molecule which is released to perform cellular functions. Consequently, the balance between the supply and the demand for ATP molecules will be an important factor in the cell's ability to function.
Oxygen is supplied to the cells by the blood and most cellular energy production is tightly coupled to oxygen. Whenever the blood flow to an organ is interrupted, a state of ischemia exists. During ischemia, cellular ATP will be consumed and usually cannot adequately be replentished in the absence of a supply of oxygen. Ischemia can exist for only a portion of an organ when the blockage of the blood supply to the organ is not total. In addition to total ischemia, or no blood flow, there are intermediate degrees of ischemia. Whenever the demand for ATP exceeds the cell's ability to produce it, cellular ATP levels will fall.
Significant ischemia occurs during most cases of open heart surgery, all episodes of coronary occlusion or heart attack, all cases of organ transplantation, certain procedures such as liver shunt operations and a variety of other situations in which either significant stress or a period of shock has compromised the functioning of one or more organs of the body. In all of these situations, cellular energy metabolism is impaired, and its restoration is critical to the recovery of organ function.
For example, much of the increased safety of heart surgery has come from improved surgical techniques which can be used when the heart is still and quiet. A quiet heart is usually produced by cooling and depriving it of its blood supply. In addition, a cardioplegic solution is frequently injected into the vessels of the heart to produce cardiac standstill and to reduce its energy demand. Although these techniques have allowed enormous progress to be made in cardiac surgery, there is still a price to be paid for this period of ischemia. The result is a period of depressed function (low cardiac output) following the operation which may or may not be tolerated by the patient. Moreover, even if it is reversed, there is evidence that scarring can occur and later failure can result. There is also an increasing emphasis on prompt reversal of the ischemia of the heart due to myocardial infarctions. The goal is to relieve the ischemia before permanent damage occurs. If the blood vessel can be unblocked by any one of a variety of methods, then the area of tissue relieved of ischemia would benefit greatly from methods to enhance its recovery. Failure of recovery from ischemia due to myocardial infarctions or to open heart surgery accounts for nearly 500,000 deaths per year in the United States.
Because of the importance of this problem, a great deal of investigation has been directed to elucidating the mechanisms responsible for the recovery of myocardial cells from ischemia. It has been found that the chemical precursors of ATP are broken down during ischemia and are not available to restore ATP levels when blood flow returns. During ischemia the cellular energy reservoir of ATP is utilized, initially producing adenosine diphosphate (ADP), and adenosine monophosphate (AMP). Further catabolism results in degradation of these products to adenosine (Ad), inosine (Ino) and hypoxanthine (Hx). With reperfusion of the organ, the recovery of ATP levels in the cell is limited because of loss of these ATP precursors. Much of the Ad, Ino and Hx have leaked out of the cells and the remaining compounds cannot easily regenerate ATP. Furthermore, endogenous synthesis of ATP precursors through the purine biosynthetic pathway, the major normal route of synthesis, proceeds slowly, is metabolically demanding, and thus limits ATP recovery.
After a period of myocardial ischemia under the conditions of clinical open heart surgery, ATP levels require about ten days to fully recover. Myocardial function has been determined to require a similar period for full return. The most sensitive aspect of myocardial function was found to be the relaxation rather than the countraction phase of the heartbeat. It is the relaxation phase, or diostole, that requires almost ten days to return to normal. When relaxation is incomplete, the heart does not fill satisfactorily and, therefore, less blood is ejected with each beat.
The theory that a reduction in the ATP recovery time could lead to improved cardiac function has lead to research aimed both at preventing the initial loss of ATP procursors from the cell and at methods for the resupply of the precursors employed in ATP biosynthesis.
Many investigators have attempted to show that specific precursors will block the fall in ATP levels or will augment ATP recovery. Adenosine, adenine, inosine, 5-amino-4-imidazolcarboxamide riboside and ribose are some of the ATP precursors that have been marginally useful in increasing ATP regeneration. Most studies were of short duration, e.g., 2 hours or less and none were longer than 24 hours. Consequently, only partial ATP recovery was found, and none accomplished the complete return of ATP levels once severe depression had been induced.
For example, H. G. Zimmer, in Science, 220, 81 (1983) reported a study in which ATP levels were shown to be maintained for 24 hours in rats which were treated with ribose after being given a toxic dose of isoproterenol and subjected to constriction of the abdominal aorta. The combined stresses of catecholamine stimulation and increased blood pressure on the heart resulted in lowered myocardial ATP levels. Zimmer's conclusion was that "the reductions in ATP and total adenine nucleotides were prevented" by this treatment. In this study the hearts were not subjected to ischemia and the ability of ribose to enhance recovery after an ATP fall had occurred was not tested.
Seifart et al. in Basic Res. Cardiol. 75, 57 (1980) studied isolated, electrically-driven guinea pig atria in which adenine and ribose were found to "inhibit the loss of cardiac adenine and pyridine nucleotides during anoxia." In this study the isolated atria were stabilized for an hour then subjected to nitrogen to cause 2 hours of anoxia, not ischemia. The addition of adenine and ribose after one hour of anoxia reduced the fall in ATP levels during anoxia. No investigation was made of the ability of adenine and ribose to restore fallen ATP levels.
Other studies have found that precursors which are relatively distant in terms of the enzymatic steps required to reform ATP appear to be less efficient in inducing ATP recovery, while other structurally closely related ATP precursors such as adenosine, can exhibit undesirable side-effects such as renal vasoconstriction.
Therefore, a need exists for a general method for the treatment of ischemic tissue which (a) rapidly restores normal cellular ATP levels and (b) maintains these levels for the time required for permanent tissue recovery from the effects of ischemia. Furthermore, a specific need exists for a method to improve the recovery of myocardial function after partial or total occlusion.